(1) Nucleotide analogs have been synthesized to use as structural probes of glutamine synthetase from Escherichia coli in order to obtain intra- and inter-subunit distances in the dodecameric enzyme. Various analogs of ATP that are substituted at the 6 or 8 position of the adenine ring have been further modified with spectrophotometric and fluorometric probes or an electron dense Pt(II) marker for electron microscopy and then introduced into active sites and/or adenylylation sites of the enzyme. (2) Glutamine synthetase active sites also have been covalently labeled and studies are in progress to locate the nucleotide binding site in the subunit primary structure. (3) Active-site ligand and metal ion interactions with mammalian glutamine synthetase are being studied to detect structural features that are similar to those of the E. coli enzyme. (4) Thermodynamic parameters for the sequential binding of active-site ligands to E. coli glutamine synthetase have been obtained by calorimetry, equilibrium binding, and fluorescent titrations. These data provide information on the modes of binding nucleotide, L-glutamate, and L-met-S-sulfoximine substrates. (5) Studies on the release of zinc ions from intact aspartate transcarbamoylase and from its regulatory subunits and the rebinding of zinc ions to regulatory subunits are in progress. The results will give new insights into the kinetic mechanisms of dissociation and asembly of this enzyme.